The broad goals are to compare the iron, reactive methionine and fatty acid binding sites of soybean lipoxygenase and reticulocyte lipoxygenase. Specific approaches include the following. (1) The products of stoichiometric additions of unsaturated fatty acids or their hydroperoxides to lipoxygenase will be determined by HPLC analysis. (2) The iron sites in several soybean lipoxygenase isozymes and in the reticulocyte enzyme will be compared by EPR. (3) The peptide containing an essential, reactive methionine will be isolated and sequenced. (4) The peptide sites of methionine oxidation and alkylation will be compared for soybean and reticulocyte proteins by peptide mapping and amino acid analysis. (5) Proximity of the reactive methionine to other regions of the protein will be examined by photocrosslinking and spin labeling. A new approach for photochemical transfer of fluorescence from methionine to nearby residues will be developed. (6) Photoreactive fatty acids will be examined by steady state kinetics to determine those with affinity for soybean lipoxygenase. Photolysis and proteolysis will be done for appropriate fatty acid derivatives to map the fatty acid binding site. (7) A study will be made of the extent of incorporation of iron isotopes into lipoxygenase by biosynthetic routes in intact reticulocytes. High levels of incorporation would allow additional physical techniques to be used in future studies of the iron environment in lipoxygenase.